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Figure 1.
Figure 1. Inhibition of LF by GM6001. (a) GM6001 inhibits
cleavage of MKKs by LF in vitro. Immunoblots show LF cleavage of
MKK-3 and MKK-1 in J774A.1 lysates in the presence of varying
concentrations of GM6001 or 10 mM o-phenanthroline, a metal
chelator. Cleavage of MKK-3 causes a mobility shift; the MKK-1
antibody is directed against the N terminus and does not react
with the cleavage product, resulting in disappearance of the
band upon cleavage. (b) GM6001 inhibits MKK-3 cleavage in lethal
toxin -treated cells. Quantified western blot analysis of MKK-3
cleavage in J774A.1 treated with lethal toxin (0.5  g
ml-1 PA with the indicated concentrations of LF) in the absence
or presence of 100 M
GM6001. (c) Protection of J774A.1 cells from lethal toxin
-mediated cell death by GM6001. Cell viability as determined by
MTT assay after lethal toxin treatment in the presence of 100
M
GM6001 or 0.2% (v/v) DMSO carrier. (d) Dose-dependent
neutralization of lethal toxin by GM6001. J774A.1 cell viability
determined by MTT assay after treatment with lethal toxin (0.5
g
ml-1 PA + 0.3 g
ml-1 LF) or PA alone (0.5 g
ml-1) in the presence of the indicated concentrations of GM6001.
(e) GM6001 protects J774A.1 cells when added subsequent to LeTx.
Cell viability is shown after treatment with PA alone (0.4 g
ml-1) or PA with LF (25 ng ml-1), with GM6001 added to 100 M
at the indicated time after toxin addition.
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